No / This article reports the structural elucidation of the Alangium alkaloid, tubulosine (1) on the basis of systematic 2D-NMR analyses (DEPT, COSY, TOCSY, NOESY, ROESY, HMQC and HMBC). The data obtained allowed the unambiguous assignment of all proton and carbon signals in 1 for the first time.

Yes / The first 1H, 13C, 31P and 11B solid state MAS NMR studies of electron- deficient carborane-containing ruthenium and osmium complexes [Ru/Os(p-cym)(1,2-dicarba-closo-dodecaborane-1,2- dithiolate)] are reported. The MAS NMR data from these 16-electron complexes are compared to those of free carborane-ligand and an 18-electron triphenylphosphine ruthenium adduct, and reveal clear spectral differences between 16- and 18-electron organometallic carborane systems in the solid state. / We thank the Swiss National Science Foundation (grant no. PA00P2-145308 to NPEB), the ERC (grant no. 247450 to PJS), EPSRC (grant no. EP/F034210/1) and EC COST Action CM1105 for support. JVH thanks EPSRC and the University of Warwick for partial funding of the solid state NMR infrastructure at Warwick, and acknowledges additional support obtained through Birmingham Science City: Innovative Uses for Advanced Materials in the Modern World (West Midlands Centre for Advanced Materials Project 2), with support from Advantage West Midlands (AWM) and partial funding by the European Regional Development Fund (ERDF).

Yes / When the 'CO-releasing molecule-3', CORM-3 (Ru(CO)3Cl(glycinate)), is dissolved in water it forms a range of ruthenium complexes. These are taken up by cells and bind to intracellular ligands, notably thiols such as cysteine and glutathione, where the Ru(II) reaches high intracellular concentrations. Here, we show that the Ru(II) ion also binds to DNA, at exposed guanosine N7 positions. It therefore has a similar cellular target to the anticancer drug cisplatin, but not identical, because Ru(II) shows no evidence of forming intramolecular crossbridges in the DNA. The reaction is slow, and with excess Ru, intermolecular DNA crossbridges are formed. The addition of CORM-3 to human colorectal cancer cells leads to strand breaks in the DNA, as assessed by the alkaline comet assay. DNA damage is inhibited by growth media containing amino acids, which bind to extracellular Ru and prevent its entry into cells. We conclude that the cytotoxicity of Ru(II) is different from that of platinum, making it a promising development target for cancer therapeutics.

Phafin2, an adaptor protein, is involved in various cellular processes, such as apoptosis, autophagy, endosomal cargo transportation, and macropinocytosis. Two domains, namely, PH and FYVE, contribute to Phafin2's cell membrane binding. Phafin2 also contains a poly aspartic acid (polyD) motif in its C-terminal region that can specifically autoinhibit the PH domain binding to membrane phosphatidylinositol 3-phosphate (PtdIns3P). Firstly, the study investigated the domain-based evolutionary pattern of PH, FYVE, and polyD motif of Phafin2 among its orthologs and Phafin2- like proteins. Using different bioinformatics tools and resources, it was concluded that the polyD motif only evolved in Phafin2 and PH- or both PH-FYVE-containing proteins of animals, highlighting the association in cellular functions that might have evolved uniquely in animals. Moreover, PH domain-free FYVE-containing proteins lack polyD motifs. Secondly, intramolecular autoregulatory and membrane binding properties of Phafin2 were studied by employing liposome co-sedimentation assay, isothermal titration calorimetry, and nuclear magnetic resonance spectroscopy. The residues Gly38, Lys45, Leu45, Lys51, Ala52, and Arg53 of the PH domain form a positively charged binding pocket that can bind the negatively charged polyD motif. The mutated Phafin2 PH domain (K51A/R53C and R53C) was unable to bind to synthetic polyD peptides, establishing the significance of those residues for the interaction between the PH domain and polyD motif. Moreover, the study also concluded that Phafin2-mediated membrane binding is not curvature-dependent. / Master of Science / Phafin2 is a protein that plays a crucial role in several important cellular functions, including cell death, recycling of cellular components, and transporting materials within cells. The protein's ability to attach to cell membranes is mainly due to two of its specific regions, the PH and FYVE domains. Additionally, Phafin2 has a section called the polyD motif that can block the PH domain from binding to specific cell membrane molecules. This study explored how these regions of Phafin2 have evolved across different species, focusing on the PH, FYVE, and polyD motifs. The findings suggest that the polyD motif is unique to Phafin2 and similar animal proteins, potentially indicating a unique role in animal cell functions. Further experiments examined how Phafin2 regulates itself and binds to cell membranes. The study identified specific amino acids in the PH domain crucial for interacting with the polyD motif. When these amino acids were altered, Phafin2 could no longer bind to synthetic polyD peptides, highlighting their importance. Finally, the research determined that Phafin2's ability to bind to membranes does not depend on the shape or curvature of the membrane.

<p dir="ltr">Biologic drugs are a key defense against many health issues. In many cases, biologic drugs are not stable in the solution state and must be lyophilized. Lyophilization in the presence of excipients increases the stability of the drug by interactions with the excipients through hydrogen bonding, which will lower the local mobility of the drug. Key threats to stability include: inhomogeneity of the drug substance and excipients, high mobility, and crystallization. Solid-state nuclear magnetic resonance spectroscopy was used to identify crystallization, assess homogeneity, and measure the local mobility of lyophilized protein and mRNA/LNP systems. </p><p dir="ltr">The impact of disaccharide type and concentration on protein stability was explored. Human serum albumin (HSA) was lyophilized with disaccharides (sucrose and/or trehalose) in different relative concentrations, and solid-state nuclear magnetic resonance spectroscopy (ssNMR) ¹H T₁ and ¹H T_1rho relaxation times were measured to determine the homogeneity of the lyophilized systems on 20-50 and 1-3 nm domains, and measure local mobility with ¹H T₁ relaxation times. HSA/sucrose systems had longer ¹H T₁ relaxation times and were slightly more stable than trehalose systems in almost all cases shown. HSA/sucrose/trehalose systems have ¹H T₁ relaxation times between the HSA/sucrose and HSA/trehalose systems and did not result in a more stable system compared to binary systems. Phase separation was evident in a sample containing relative concentrations of 10% HSA and 90% trehalose, suggesting trehalose crystallization during lyophilization. Under these stability conditions, a ¹H T₁ relaxation time below 1.5 s correlated with an unstable sample, regardless of disaccharide(s) used.</p><p dir="ltr">The effect of mannitol on protein stability was studied. Human serum albumin was lyophilized in binary systems with mannitol, and in ternary systems with sucrose or trehalose and mannitol. The monomer content of the HSA was monitored over 36 weeks of storage at 50 C. The amount of mannitol in the system dictated the ability of mannitol to crystallize, and the polymorph that mannitol crystallized into. In HSA/mannitol systems, mannitol crystallization caused inhomogeneity of the matrix, determined by ¹H T_1rho relaxation times. Adding a disaccharide to the matrix, however, increased the homogeneity of the matrix. Addition of mannitol to a HSA/disaccharide matrix resulted in less stability at similar HSA:disaccharide ratios.</p><p dir="ltr">The impact of storage temperature on protein stability was investigated. Human serum albumin was lyophilized with sucrose or trehalose in histidine, phosphate, or citrate buffer. ¹H T₁ relaxation times were measured by ssNMR and were used to assess the homogeneity and mobility of the samples after zero, six, and twelve months at different temperatures. The mobility of the samples decreased after 6 and 12 months storage at elevated temperatures, consistent with structural relaxation of the amorphous disaccharide matrix. Formulations with sucrose had lower mobility and greater stability than formulations with trehalose.</p><p dir="ltr">The effect of an RF-assisted lyophilization method on homogeneity, mobility, stability, and moisture content was explored. This method, utilizing 18 GHz microwave frequency to accelerate the lyophilization cycle, resulted in equivalent or better stability for attenuated live virus or protein formulations, respectively. ssNMR showed comparable amounts of homogeneity in the formulations, however mobility of the samples produced by RF-assisted lyophilization was slightly higher.</p><p dir="ltr">A lyophilized mRNA/LNP formulation was prepared. Disaccharide type, disaccharide concentration, and freezing rate were found to alter critical quality attributes of the system. When mRNA/LNP formulations were stored at 4 C, solution formulations outperformed lyophilized formulations for at least 6 months. When mRNA/LNP formulations were stored at room temperature, solution formulations were superior for the first three months, however lyophilized formulations outperformed solution formulations after 6 months, with less growth in particle size and less loss of encapsulation efficiency. ssNMR was used to assess the interactions between the formulation components.</p>

The diffusion behavior of guest molecules introduced in porous materials has been studied. Diffusion studies in such porous materials may help for elucidating the structural properties, transport mechanism and/or surface barriers of the zeolite structure. The focus of this work is on diffusion in nanoporous materials with complex pore spaces. First a short introduction in the basics of diffusion and the PFG NMR technique (Pulsed Field Gradient Nuclear Magnetic Resonance) is described. In the following two chapters the diffusion in hierarchical pore spaces or, to be more precise, zeolites with generated mesopores, which traverse the microporous bulk phase, are investigated. The hierarchical pore spaces consists in the first case of micro- and mesopores and in the second case of micro-, meso- and macropores. The diffusion behavior in these materials has been investigated revealing diffusion acceleration in the mesoporous samples, as compared to the purely microporous material. In the next chapter the diffusion behavior in glass samples with different porosity and their complementary pore space is investigated. Diffusion with full loaded pore spaces and surface diffusion, where the molecules were only able to diffuse along the pore walls, has been explored. The aim was to find out to what extent the diffusion in two complementary pore spaces is correlated. In the last chapter, the effect of an inorganic binder on the transport in zeolite pellets has been studied. First the diffusion behavior in binderless zeolite beads in comparison with the zeolite powder employed for their production has been explored. The particular interest was to find out up to which extent the diffusion patterns observed with the powder samples could again be recognized in the beads. In a second study the transport characteristics within binderless molecular sieves have been investigated, with the purpose to reveal differences in the diffusion behavior in comparison with their binder-containing counterparts.

No description available.

No description available.

Specific ion effects, identified for more than hundred years, play an important role in a wide range of phenomena and applications. Several mechanisms such as direct ion interaction with molecules have been suggested to explain these effects, but quantitative experimental evidence remains scarce. Electrophoretic NMR (eNMR) has been emerging as a very powerful tool for studying molecular association and ionic transport in a variety of systems. Yet its potential in studying specific ion effect has been unexplored. In this thesis, eNMR was in part developed further as an analytical method and was in part used as one of the main techniques to study ions interacting with macromolecules in aqueous and non-aqueous solutions. The complexation of a large group of cations with poly ethylene oxide (PEO) in methanol was studied with eNMR. The binding of monovalent ions was demonstrated not to follow the Hofmeister order; multivalent cations except barium all showed negligible complexation. As a unifying feature, only cations with surface charge density below a threshold value were able to bind suggesting that ion solvation is critical. The binding mechanism was examined in greater detail for K+ and Ba2+ with oligomeric PEO of different chain lengths. Those two cations exhibited different binding mechanisms. K+ was found to bind to PEO by having at least 6 repeating units wrap around it while retaining the polymer flexibility. On the other hand, Ba2+ (and, to some extent, (BaAnion)+) needs a slightly shorter section to bind, but the molecular dynamics at the binding site slow considerably. The binding of anions with poly (N-isopropylacrylamide) in water was quantified at low salt concentration with eNMR and the binding affinity, though very weak, followed the Hofmeister order. This result indicates the non-electrostatic nature of this specific ion effects. The increase of binding strength with salt concentration is well described by a Langmuir isotherm. The specific ion binding to a protein, bovine serum albumin (BSA), was also studied at pH values where BSA has either net positive and negative charges. Our results show that anions have the same binding affinity irrespective of the surface charge while the binding strength of cations is reversed with the change in net surface charge. This indicates different binding mechanisms for cations and anions. The ionization of cellobiose in alkaline solutions was measured quantitatively by eNMR. The results show a two-step deprotonation process with increasing alkaline strength. Supported by results from 1H-13C HSQC NMR and MD simulation, ionization was proposed to be responsible for the improved solubility of cellulose in alkaline solution. eNMR was also used to characterize the effective charge of tetramethylammonium ions in a variety of solvents. In solvents of high polarity, the results agree well with predictions based on Onsager’s limiting law but for nonpolar solvents deviations were found that were attributed to ion pair formation. / <p>QC 20170216</p>

This thesis deals with black carbon, its characteristic features and with its occurrence in chernozemic soils. In particular, this thesis deals with methods, which can study presence of black carbon in soils. The presence of black carbon and the whole composition of soil organic matter of three chernozemic soils in Czechia (Zeměchy, Tursko, Syrovice) is examined by 13 C NMR spectroscopy. Anthracological analysis of charcoal from fossil chernozems of Zemechy loess ravine deals with pedogenesis of chernozems and development of Quaternary vegetation in Central Europe. Coniferous tree species of Pinus sp., Pinus cf. cembra, Larix/Picea, Juniperus a Vaccicium, i.e. cold- and drought-tolerant taxa, were identified by anthracological analysis of soils of Zemechy loess ravine. The identified species suggest that the landscape around Zemechy was probably formed by parkland taiga. According to 13 C NMR spectroscopy, soil organic matter of fossil chernozem of Zemechy loess ravine consists particularly of alkyl and O-alkyl carbon. Aromatic carbon is also significant. O-alkyl carbon is the most important in the recent chernozems of Tursko and Syrovice. Aromatic carbon has the smallest proportion in both chernozems. The presence of aromatic carbon in chernozem of Tursko is the smallest of all analyzed soils. The...